Fig. 3

Multiple Ser/Thr (S/T)-rich motifs in ATF2 are required for SPOP-mediated ATF2 degradation. a Diagram showing wild-type ATF2 proteins and S/T rich motif-deleted mutants. The S/T rich motif is depicted in red. b Western blot of WCL of 293 T cells transfected with indicated plasmids. c Western blot of WCL and co-IP samples of anti-FLAG antibody obtained from 293 T cells transfected with indicated plasmids. d and e Western blots of WCL from 293 T cells transfected with the indicated constructs, treated with 50 μg/ml cycloheximide (CHX) and harvested at different time points. e Quantification of the western blots carried out in d. At each time point, the intensity of ATF2 protein was normalized to the intensity of actin and then to the value at 0 h. f Western blots of the products of in vivo ubiquitination assays performed using cell lysate from 293 T cells transfected with the indicated plasmids and treated with 20 μM MG132 for 8 h